One promising new therapy for Parkinson's Disease (PD) involves the replacement of degenerated nigrostriatal neurons with those derived from transplanted fetal mesencephalic tissue. Although this approach has often yielded remarkable recovery of function in rats and monkeys, results in clinical trials with PD patients have been less consistent. At issue, is the relative inability to standardize a number of critical factors in human fetal transplants, including the age, type, number and integrity of cells being grafted. Consequently, finding more reliable sources of dopaminergic (DA) tissue for transplantation has become increasingly important. One direction has been to search for a line of readily available, well-characterized continually self-renewing stem or precursor cells that possess the capacity to differentiate, ideally spontaneously and with the need for little manipulation, into DA neurons, thus providing an inexhaustible and uniform source of replacement tissue. Towards this end, our preliminary findings demonstrate that grafts of embryonic mouse neural stem cells (NSCs) of the C17.2 cell can differentiate exclusively into neurons, which in a majority of cases, can express DA traits when cells are transplanted into the brain of a Parkinsonian rat. In addition, in preliminary studies using stem cells from adult human bone marrow (MSCs), we have found that nearly 100 percent of MSCs will convert into process- bearing, beta-tubulin III+ neuronal-like cells after only 1-2 hours of incubation with specific differentiation factors. If these cells also exhibit the same capacity as NSCs to respond to appropriate DA differentiation cues in vivo, patients could provide their own source of stem cells for autologous grafts in PD. Using NSC and MSC stem cell models and a multidisciplinary approach, our specific goals for this proposal are threefold: 1) Identify the conditions that promote the stable appearance of a postmitotic differentiated DA phenotype in stem cells grown in culture; 2) Identify those factors which promote the differentiation of a DA phenotype in transplanted stem cells and 3) Determine whether the DA phenotype in transplanted stem cells is stable and long lasting, and whether, it can produce functional recovery of motor deficits in a rat model of PD. The ultimate goal of this research program is a fuller understanding of the cellular and molecular processes regulating the differentiation of DA traits in stem cells and apply that knowledge to transplantation strategies for the treatment of Parkinson's Disease.